1. Field of the Invention
The present invention relates to capillary electrophoresis, and particularly an improved capillary tube for use to conduct capillary electrophoresis analysis.
2. Description of Related Art
Synthetic fused silica capillary tubing is widely used in the separation sciences, including the popular fields of Gas Chromatography, Capillary Liquid Chromatography, and Capillary Electrophoresis (CE). From its advent in the late 1970's, steady advances in product quality have emerged. Light guiding fused silica capillary tubing by applying soft fluoropolymer coating is the latest step in this continuing development. [See, Macomber Joe, Nelson Gary Light Guiding Fused Silica Capillary Tubing, As printed in LCGC, The APPLICATION NOTEBOOK—June 2002, Page 48.] Improved purity of the synthetic fused silica, more durable external coatings, and tightening of specification tolerances are cornerstones in capillary tubing advancements. Researchers have successfully used capillary tubing filled with a high refractive index liquid to form an optical wave-guide. In one specific application, a low refractive index, external soft fluoropolymer coated capillary was employed in a Raman spectroscopy device. [See, D. Che and S Liu, Long Capillary Waveguide Raman Cell, U.S. Pat. No. 5,604,587, 1997.] Although a viable product for light guiding applications, soft fluoropolymer coated capillary has never achieved the low cost or durability desired for the large volume disposable multi-channel capillary gel cartridges of the capillary electrophoresis type instrument.
High-performance capillary electrophoresis (HPCE) now represents a set of powerful electromigration techniques whose impact has been felt in virtually all areas of biochemical analysis. [See, Novotny, M. V.; Sudor, J. Electrophoresis 1993, 14, 373-389; Novotny, M. V. High Performance Capillary Electrophoresis, Theory Techniques and Applications, John Wiley & Sons: New York, 1998, Chapter 21, pp 729-765; Novotny, M. V. Methods Enzymol. 1996, 270, 101-133; Stefansson, M., Novotny, M. V. Techniques In Glycobiology, Marcel Dekker, New York, 1997, Chapter 26, pp 409-430.] HPCE is a micro fluidic approach to gel electrophoresis, whose greatest advantage is its diverse range of applications. CE technology with fluorescence based detection is commonly accepted by the biotechnology industry as a reliable, high resolution and highly sensitive detection tool. [See, Guttman, A. and Cook, N. Anal. Chem. 1991 63, 203 8-2042.]
The existing commercial CE systems with laser-induced fluorescence (LIF) detection mechanism that use multiple capillaries/channels for high-throughput applications are complicated in design and operation of the instrument. These systems utilize fused silica capillaries with polyimide coated jacket and a clear window area (i.e., with polyimide jacket removed at the window area) at the detection zone. Soft fluoropolymer coated capillaries have not been used in such multi-channel applications. The polyimide coating (jacket) provides strength and prevents the small OD glass capillaries from breakage. For fluorescence based detection, the polyimide jacket at the detection zone of the capillary must be removed. It is essential that the detection zone of the capillary tube to be completely free of any polyimide material or particulates to provide a clear optical path through the fused silica glass for excitation and direct emission detection/collection of fluorescence signal from the samples (bio-molecules) that are migrating or flowing inside the ID of the tubes.
To remove the polyimide jacket to provide a clear optical widow at the detection zone of the capillary for fluorescence detection, the polyimide jacket could be burned off by applying heat (using hot electric coils), acid etching, mechanical type scraping or removal (e.g., using a razor blade), or the jacket could be removed by photo-etching or photo-ablation techniques using different wavelengths and/or types of lasers. This process is very labor intensive and requires a careful examination of the glass surface under high magnification after the removal process to make sure the window (glass surface) is completely free of the polyimide jacket or any other particulates. Polyimide also fluoresces under visible light, which introduces further complications in the detection path. The removal of the polyimide jacket at the detection zone (e.g., a 3-5 mm long section) also poses weak joint exposing the bare capillary for possible micro-cracks or complete breakage. Excessive heat or mechanical stresses applied at the detection zone during the removal of the coated materials/jacket also produces micro-cracks in the OD and ID of the fused silica tube, which are not visible or detectable under the high power magnification, which also can increase the background fluorescence affecting the overall Limit of Detection (LOD) in CE type instruments, such as DNA fragment analysis type CE instruments.